A human disease model for the study of genomic imprinting. Prader- Willi syndrome (PWS) is a complex neurodevelopmental disorder caused by the inactivation of deletion of imprinted, paternally expressed genes within 4 Mbp of chromosome band 15q11.2. The overall goals of this project are to determine the mechanisms of imprinting in the PWS region and to elucidate the genetic and pathophysiologic pathways that lead to the PWS phenotype that involves metabolic and behavioral changes such as growth deficiency and lack of appetite control. A recently discovered novel paternally expressed imprinted gene cluster in the PWS deletion region (PWCR1 in human and Pwcr1 in mouse) encodes small nucleolar RNAs (snoRNAs) with short conserved sequence elements, called C and D boxes (de Los Santos et al. Amer. J. Hum. Genet. 67:1067, 2000). PWCR1 and Pwcr1 are the first mammalian snoRNA genes that show imprinted expression. C/D box snoRNAs are conserved in all organisms and usually serve to direct the site-specific methylation of the ribose 2-hydroxyl group of specific nucleotides in rRNA., PWCR1/Pwcr1, belong to a novel class of snoRNAs, found mostly in brain, in which the sequences that would basepair with the target RNA are not complementary to any known rRNA or snRNA sequences. It is hypothesized that the modification target(s) of PWCR1snoRNAs are brain-specific mRNAs whose processes, such as alternative splicing, methylation or other modification, or intranuclear trafficking is controlled by binding to snoRNA-containing particles; and that lack of PWCR1 snoRNAs contributes to the hypothalamic dysfunction that causes the physical and behavioral manifestation of the PWS phenotype. The proposed work will characterize the organization, transcription, processing, localization and function of this novel kind of snoRNAs. Experimental strategies include the mining of the human genome for organization of the PWCR1snoRNA gene cluster, for a putative "host" gene for the PWCR1snoRNA genes and for candidate RNA modification targets. The expression patterns of putative target RNAs will be studied in the brain from PWS patients and in a PWS mouse model in which the Pwcr1 snoRNA cluster is deleted. PWCR1 snoRNAs will be tested for function by primer extension studies to detect 2'-O-methylation of synthetic complementary target RNAs. The work has the potential for discovery of novel RNA molecules that are modified as a result of a complex formation between PCR1snboRNPs and target RNA, and thus, for major breakthroughs in understanding the role of mRNA modification, and for innovation of new diagnostic and treatment modalities.